Many cellular processes are mediated by the regulation of small GTPase signaling cascades. The most widely studied of these signaling pathways are those of the Ras superfamily, of which there are four subfamilies; Ras, Rho/Rac, Rab and ARF.
Within these pathways, however, a common theme of regulation exists. GTPases cycle between active and inactive forms. When bound to GDP they are inactive, and when bound to GTP they are active and able to transduce signals to effector molecules which will ultimately transmit the signal further downstream to the next protein.
Since the GTPase family of proteins is involved in a diverse and complex system of signaling cascades, the on/off switch of GTPases is highly regulated. There are two types of proteins that regulate the activity of GTPases. Those that activate, or enhance, the binding of GTP are known as guanine nucleotide exchange factors (GEFs). Those that enhance the GTPase's intrinsic ability to hydrolyze GTP to GDP, thereby inactivating the GTPase, are called GTPase activating proteins (GAPS).
RIP-1 (also known as RalBP1 and RLIP) is a GTPase activating protein (GAP) that was identified based on its ability to bind to the GTP-bound form of a GTPase known as Ral. However, it was shown that this binding does not activate Ral's intrinsic GTPase activity (Matsubara et al., FEBS Lett., 1997, 410, 169-174). It is therefore believed that RIP-1 is an effector, or downstream target, of Ral. It was further suggested that the relationship between Ral and RIP-1 is necessary to translocate RIP-1 to the membrane (the location of the Ral protein) where it can subsequently interact with the RIP-1 effectors, CDC42 and Rac1 (Matsubara et al., FEBS Lett., 1997, 410, 169-174).
Upon binding CDC42 or Rac1, RIP-1 activates the intrinsic GTPase activity of these proteins causing them to hydrolyze their bound GTP, thereby attenuating the signals from them. Furthermore, these proteins have been shown to regulate the formation of filopodia and lamellipodia, respectively. Taken together, these studies indicate a role for RIP-1 in cell morphology (Feig et al., Trends Biochem. Sci., 1996, 21, 438-441).
Moreover, due to the association of RIP-1 with these regulators of cytoskeletal organization, altered RIP-1 regulation is likely to be involved in the formation or progression to diseased states. Cellular transformation and acquisition of the metastatic phenotype are the two main changes normal cells undergo during the progression to cancer. CDC42 and Rac1 are known to be involved in Ras-mediated transformation of cells. In addition, they have also been shown to activate kinase cascades that lead to gene activation from the serum response factor (Feig et al., Trends Biochem. Sci., 1996, 21, 438-441). Thus RIP-1 may play a role as an upstream regulator of these aberrant cellular processes.
Currently, there are no known therapeutic agents which effectively inhibit the synthesis of RIP-1. To date, strategies aimed at inhibiting RIP-1 function have involved the use of enzymes which inhibit the posttranslational modification of Ral. This method interferes with the localization of Ral to the membrane which eliminates the recruitment of RIP-1 to the membrane, and subsequently blocks the interaction of RIP-1 with its effectors, CDC42 and Rac1.
However, these strategies are not specific to Ral, as many proteins undergo similar posttranslational modifications. Consequently, there remains a long felt need for additional agents capable of effectively inhibiting RIP-1 function.